S254: CD4+ T CELL-DERIVED IL21 REGULATES STEM CELL FATE IN ACUTE MYELOID LEUKEMIA
Authors: V. Rubino
Journal: HemaSphere (2022)

Background: Acute myeloid leukemia (AML) is characterized by poor prognosis for patients, due to the very high rate of relapse, which occurs even after chemotherapy-induced complete remission. Therapy-insensitive AML cells, so-called leukemia stem cells (LSCs), are thought to be the main driver of relapse. In fact, high LSCs number and active LSC self-renewal programs are related to poor therapy response and adverse outcome in AML. Therefore, eliminating LSCs is an urgent, yet unmet, medical need. LSCs properties are crucially regulated and maintained also through interactions with the bone marrow microenvironment, which includes immune cells. Despite evidences of mutual interactions between AML cells and immune cells, their interplay is still poorly understood. A better understanding of these interactions might provide novel methods to effectively eradicate LSCs and achieve durable AML cure. Aims: The aim of this study was to investigate the role of IL21/IL21R signaling in the so far unexplored context of AML. Our aims involved identification of IL21 source in murine and human AML and elucidation of the mechanisms how IL21/IL21R signaling regulates LSCs properties. Methods: We used a combination of flow cytometry, ELISA and qRT-PCR to determine protein and mRNA expression of both IL21 and IL21R in primary AML samples and analyzed the correlation between IL21 serum levels with patients’ survival and complete remission rate. Colony-forming assays and short-term cultures were used to determine the in vitro effect of IL21 on primary normal and leukemic stem cells. In addition, patient-derived xenografts were employed to study IL21 effect on stem cell frequency and AML engraftment capacity. We used various syngeneic murine AML models (e.g. MLL/AF9 and MLL/ENL) to study the functional role of IL21/IL21R signaling in vivo. We performed RNA-Seq, together with analysis of symmetric cell division, expression of differentiation markers, NF-kB signaling, stem cell frequencies by limiting-dilution secondary transplantation experiments to investigate the mechanisms how IL21 regulates LSCs. Results: In human AML, serum IL21 was identified as an independent positive prognostic biomarker for overall survival (OS). Furthermore, high dose chemotherapy was more effective in patients with high levels of IL21 at diagnosis, as illustrated by a significantly higher rate of complete remission (CR rate: 81 vs 59 %) and prolonged OS (median survival: 915 vs. 364 days, HR: 1.8). Functionally, IL21 inhibited cell growth and clonogenic potential of primary AML stem and progenitor cells but not normal hematopietic stem cells (HSCs). In murine AML models, IL21/IL21R signaling in LSCs reduced stem cell frequency, prolonged survival of the mice and resulted in the downregulation of stemness-related and the up-regulation of differentiation-promoting pathways. In both human and murine AML, CD4+ T cells were identified as the main source of IL21. Image:Summary/Conclusion: In this work, we have identified IL21/IL21R signaling pathway as an important regulator of cell fate in human and murine LSCs, but not HSCs. We found that IL21 is a positive prognostic marker for OS and that higher serum IL21 levels correlate with better survival and higher rate of complete remission in patients that undergo chemotherapy. Our findings therefore suggest that promoting IL21/IL21R signaling on LSCs may be a novel approach to decrease stemness and increase differentiation in AML.